Interspecific fusion of sea urchin eggs: Surface events and cytoplasmic mixing

Abstract

Here we describe a new method of fusing fertilized and unfertilized sea urchin eggs. After removal of materials peripheral to the plasma membranes, the plycation polyarginine is used to induce the eggs to adhere to one another. The subsequent incubation of the eggs in an artificial sea water with a slightly lower osmolarity and a higher calcium content than normal sea water results in successful fusion. We demonstrate with scanning electron microscopy (SEM) that our artificial sea water serves to smooth the plasma membranes—a characteristic which we find to be a requirement in order for fusion to occur. We also demonstrate that our fusion procedure can be used to identify and follow the development of hybrids composed of eggs of different histories. Using species whose cytoplasms are distinguishable by light microscopy, Strongylocentrotus purpuratus and Lytechlnus pictus, we consider the mixing of cytoplasms using yolk granules as markers. We find that if two unfertilized eggs are fused, the species-specific yolk particles are free to mix. In contrast, we observe no mixing of the yolk particles in either fertilized-unfertilized or fertilized-fertilized interspecies hybrids until the time of mitosis. At that time, the yolk particles are free to move around the region containing the mitotic figure(s). However, mixing of the cytoplasmic yolk granules is never completed during this brief period. Even after cytokinesis takes place, one can see the segregation of the yolk granules in the cytoplasms of the two species.